The aim of this research is to obtain basic information on the respiratory chain enzyme system of mitochondrial inner membrane enabling conclusions on carriers, their sequence, actions, and mechanisms of electron (hydrogen) transfer which eventually leads to energy coupling. The methods planned will be mainly by sequential resolution, enzyme identification and finally systematic reconstitution. Among others, the resolved enzymes and components are first purified (crystallized or crystallized in "2-dimensional" way, if possible, then examined by electron microscope and image reconstruction) and rigorously characterized by various chemical, physico-chemical, and enzymic techniques including amino acid sequence determination. It must be emphasized that reconstitution will be used as a most reliable technique (at present at least) to ascertain no denaturation or changes occurred in the extensive purification of components. An ultimate goal, not necessarily the eventual goal, in our first phase is reconstitution of the complete electron transport chain which constitutes a part of bioenergetic apparatus with all functional manifestations of the mitochondrion. This proposal differs from NIH-GM16767 in the emphasis of studies of non-cytochrome containing carriers in the present project.